Expanded foam for delivery of functional ingredients

ABSTRACT

The present disclosure provides delivery systems for functional ingredients, such as drugs, nutritional supplements, botanicals, and vitamins. The delivery systems comprise an ingestible matrix within which the functional ingredient(s) are substantially uniformly and completely dispersed. The matrix comprises foams derived by physical shear of egg white to achieve amplified available dose and improved release characteristics of functional ingredients at skin and mucosal surfaces for healthcare purposes. The disclosure also provides methods of preparing and using the delivery systems.

FIELD OF THE DISCLOSURE

The present disclosure relates to the field of health components orfunctional ingredients including medicaments and nutrients and a systemfor delivering the health components or functional ingredients.

BACKGROUND OF THE DISCLOSURE

Functional ingredients including drug products such as antibiotics,antiseptics and topical disinfectants designed to treat or preventinfectious disease and nutritionals including vitamins, amino acids andessential fatty acids designed to promote health and well-being inhumans and animals have grown in popularity, as evidenced by thetremendous growth in the industry involved in their manufacture,production, and distribution.

Among the various classes of functional ingredients many arewater-soluble, some are only soluble in oil while a few are amphipathicdisplaying partial solubility in oil and water in the manner of asurfactant.

The solubility of any functional ingredient greatly influences theappropriate manner of its delivery to a human or animal body,particularly oral applications where the functional ingredient isintended to affect a response on or in the gastro-intestinal tract andin topical and mucosal applications where the functional ingredient isintended to affect a response on or in the skin or mucosal membranes andby extension on structures emanating from these surfaces such as hair,nails and teeth.

Formulations of poorly soluble functional ingredients normally containexcipients designed to facilitate solubility, absorption and/orresidence time on an intended surface. The available dose of anyfunctional ingredient in such formulations is also limited by the amountof formulation that can be applied to the intended surface and themigration or release of the functional out of the formulation and ontoor into the intended surface.

Several different delivery systems have been developed to attempt toimprove methods of delivering various supplements or functionalingredients. For example, a number of encapsulated formulations havebeen developed which encapsulate or retain functional ingredients invarious glassy, sintered, or chewy confectionery-type matrixes. Ingeneral, the confectionery serves as a solid continuous matrix for theactive ingredient or supplement. The active ingredient is deliveredaccording to the dissolution rate of the confectionery matrix, whichconfers a solid taste in the mouth. Crushing the confectionery is asolution for the consumer to speed up the release of the activeingredient but this solution may be undesirable as dental problems mayarise and/or the release rate of the active ingredient incorporatedtherein may no longer be optimal. Depending upon the method ofmanufacturing the confectionery matrix, the active ingredient may sufferfrom deterioration or damage due to heat and/or mechanical stresses inthe manufacturing process.

Often, high deterioration rates due to strong processing conditions arecompensated for by overdosing of the active ingredient in theconfectionery matrix, however, this is a costly method resulting in thewastage of a lot of the active ingredient. The “solid” taste a pressedtablet or glassy matrix may provide in the mouth may also be consideredas not very attractive in the context of delivering active ingredients,especially if taste is unpalatable.

Accordingly, it is desirable to provide a system for efficientlydelivering the functional ingredient(s) with improved solubility andrelease characteristics in particular for amphipathic and oil solublefunctional ingredients.

SUMMARY OF DISCLOSURE

In accordance with an aspect of the present disclosure, there isprovided a delivery system for one or more functional ingredient(s)wherein the delivery system represents an expanded foam matrix whereinthe one or more functional ingredient(s) are substantially uniformlydispersed, said matrix comprising:

i) an egg white component comprising between 1-50% proteinconcentration;

ii) one or more heat resistant and/or heat sensitive gelling agents;

iii) a pH regulator;

iv) one or more plasticizers and/or humectants; and

v) one or more source of water, wherein said delivery system is a solidat room temperature.

In another aspect the present disclosure, there is provided a use of adelivery system for one or more functional ingredient(s) for oraladministration to an animal in need thereof.

In embodiments, the present disclosure provides methods for improvingoral health of companion animals.

In an embodiment the present disclosure provides a method of maintainingor improving oral health in a subject in need thereof, the methodcomprising administering to the subject an effective amount of an oralantimicrobial composition, wherein the oral antimicrobial compositioncomprises: (a) one or more saturated or unsaturated free fatty acids ora pharmaceutically acceptable salt thereof; and (b) one or moredelipidised membrane lipids, as emulsifying agent for the free fattyacid(s) or the salt thereof.

In embodiments, the present disclosure provides methods for oraldelivery of a health related composition comprises one or more healthcomponents for use in oral health, joint health and mobility,cardiovascular health, bone health, skin health, gut health,anti-stress/calming or other behavioral conditions, anti-parasiticidessuch as anti-flea or anti-tick or vaccines to a companion animal.

Additional variations and advantages of this disclosure will becomeapparent from the detailed description of this disclosure taken inconjunction with the accompanying Examples.

DETAILED DESCRIPTION

It is noted that in this disclosure and particularly in the claims,terms such as “comprises”, “comprised”, “comprising” and the like canmean “includes”, “included”, “including”, and the like; and that termssuch as “consisting essentially of” and “consists essentially of” allowfor elements not explicitly recited, but exclude elements that are foundin the prior art or that affect a basic or novel characteristic of thedisclosure.

Unless otherwise noted, technical terms are used according toconventional usage. Definitions of common terms in molecular biology maybe found in Benjamin Lewin, Genes V. published by Oxford UniversityPress, 1994 (ISBN 0-19-854287-9); Kendrew et al. (eds.), TheEncyclopedia of Molecular Biology, published by Blackwell Science Ltd.,1994 (ISBN 0-632-02182-9); and Robert A. Meyers (ed.), Molecular Biologyand Biotechnology: A Comprehensive Desk Reference, published by VCHPublishers, Inc., 1995 (ISBN 1-56081-569-8).

As used above, and throughout the description of the disclosure, thefollowing terms, unless otherwise indicated, shall be understood to havethe following meanings: The singular terms “a,” “an,” and “the” includeplural referents unless context clearly indicates otherwise. Similarly,the word “or” is intended to include “and” unless the context clearlyindicate otherwise. The word “or” means any one member of a particularlist and includes any combination of members of that list.

It will also be understood that, although the terms first, second, etc.may be used herein to describe various elements, these elements shouldnot be limited by these terms. These terms are only used to distinguishone element from another. For example, a first gesture could be termed asecond gesture, and, similarly, a second gesture could be termed a firstgesture, without departing from the scope of the present disclosure. Allmethods or processes described herein can be performed in any suitableorder unless otherwise indicated herein or otherwise clearlycontradicted by context.

The term “about,” as used herein, means approximately, in the region of,roughly, or around. When the term “about” is used in conjunction with anumerical range, it modifies that range by extending the boundariesabove and below the numerical values set forth. In general, the term“about” is used herein to modify a numerical value above and below thestated value by a variance of 10%. In one aspect, the term “about” meansplus or minus 10% of the numerical value of the number with which it isbeing used. Therefore, about 50% means in the range of 45%-55%.

Numerical ranges recited herein by endpoints include all numbers andfractions subsumed within that range (e.g., 1 to 5 includes 1, 1.5, 2,2.75, 3, 3.90, 4, and 5). It is also to be understood that all numbersand fractions thereof are presumed to be modified by the term “about.”

The terms “subject,” “patient,” “user,” and “individual” are usedinterchangeably herein, to refer to a human or an animal.

The terms “animal” and “companion animal” are used herein to include allmammals, birds and fish. The animal as used herein may be selected fromthe group consisting of equine (e.g., horse), canine (e.g., dogs,wolves, foxes, coyotes, jackals), feline (e.g., lions, tigers, domesticcats, wild cats, other big cats, and other felines including cheetahsand lynx), bovine (e.g., cattle), swine (e.g., pig), ovine (e.g., sheep,goats, lamas, bison), avian (e.g., chicken, duck, goose, turkey, quail,pheasant, parrot, finches, hawk, crow, ostrich, emu and cassowary),primate (e.g., prosimian, tarsier, monkey, gibbon, ape), humans, andfish.

The use of any and all examples, or exemplary language (e.g., “such as”)provided herein, is intended merely to better illuminate the disclosureand does not pose a limitation on the scope of the disclosure unlessotherwise indicated. No language in the specification should beconstrued as indicating any element is essential to the practice of thedisclosure unless as much is explicitly stated.

The term “carrier” refers to a diluent, adjuvant, excipient, or vehiclewith which the active/functional component is administered. Suchpharmaceutical carriers can be sterile liquids, such as water and oils,including those of animal, vegetable or synthetic origin, such as peanutoil, soybean oil, palm oil, mineral oil, sesame oil and the like. Wateror aqueous solution saline solutions and aqueous dextrose and glycerolsolutions are preferably employed as carriers, particularly forinjectable solutions. Alternatively, the carrier can be a solid dosageform carrier, including but not limited to one or more of a binder (forcompressed pills), a glidant or lubricant, an encapsulating agent, aflavorant, and a colorant. Suitable pharmaceutical carriers aredescribed in “Remington's Pharmaceutical Sciences” by E. W. Martin (MackPublishing Co., Easton, Pa.); Gennaro, A. R., Remington: The Science andPractice of Pharmacy, (Lippincott, Williams and Wilkins); Liberman, etal., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y.;and Kibbe, et al., Eds., Handbook of Pharmaceutical Excipients, AmericanPharmaceutical Association, Washington.

The term “effective amount” or “effective dose” as used herein refers tothat amount of a health component that is known in the art to confer ahealth benefit; wherein the effective amount in the composition is highenough to provide the desired effect or benefit to the subject, yet lowenough to avoid adverse effects such as toxicity, irritation, orallergic response, commensurate with a reasonable benefit/risk ratiowhen used in the manner of the present disclosure. Such effectiveamounts are readily ascertained by one of ordinary skill in the art andwill vary with such factors as the specific health component used, theparticular condition being treated, the age and general health of thesubject, the duration of the treatment, the nature of concurrent therapy(if any), the specific dosage form to be used, the carrier employed, thesolubility of the dose form, and the particular dosing regimen.

The terms “health component,” “functional component,” “healthingredient” and “functional ingredient” are used interchangeably hereinas used herein refers to components or ingredients which promote healthand well-being, prevent disease, or enhance well-being includes drugproducts such as antibiotics, antiseptics and topical disinfectantsdesigned to treat or prevent infectious disease, antioxidants,phytochemicals, hormones, vitamins such as vitamins A, B1, B2, B6, B12;C, D, E, K, pantothenate, folic acid, pro-vitamins, minerals such ascalcium, selenium, magnesium salts, available iron, and iron salts,microorganisms such as bacteria, such as live lactobacilli, fungi, andyeast, prebiotics, probiotics, trace elements, essential and/or highlyunsaturated fatty acids such as omega-3 fatty acids, and mid-chaintriglycerides, nutritional supplements, enzymes such as amylases,proteases, lipases, pectinases, cellulases, hemicellulases,pentosanases, xylanases, and phytases, pigments, oligopeptides,dipeptides, and amino acids, and mixtures thereof.

The term “prebiotics” as used herein refers to a “non-digestible foodcomponents that beneficially affect the host by selectively stimulatingthe growth and/or activity of one or a limited number of bacteria in thecolon that can improve the health of the host” for example, in Gibson,G. R. & Roberfroid, M. B., Dietary Modulation of the Human ColonicMicrobiota-Introducing the Concept of Probiotics, J. Nutr. 125:1401-1412(1995). Such prebiotics may be naturally-occurring, synthetic, ordeveloped through the genetic manipulation of organisms and/or plants,whether such new source is now known or developed later. Prebioticsuseful in the present disclosure may include oligosaccharides,polysaccharides, and other prebiotics that contain fructose, xylose,soya, galactose, glucose and mannose, for example, in Ramirez-Farias etal., Br J Nutr (2008)4:1-10; Pool-Zobel and Sauer, J Nutr (2007),137:2580 S-2584S. More specifically, prebiotics useful in the presentdisclosure may include lactulose, lactosucrose, raffinose,gluco-oligosaccharide, inulin, polydextrose, polydextrose powder,fructo-oligosaccharide, isomalto-oligosaccharide, soybeanoligosaccharides, lactosucrose, xylo-oligosacchairde,chito-oligosaccharide, mannan oligosaccharides or mannooligosaccharides(MOS), aribino-oligosaccharide, siallyl-oligosaccharide,fuco-oligosaccharide, galacto-oligosaccharide, andgentio-oligosaccharides. Furthermore, prebiotics useful in the presentdisclosure include molecules such as beta-methyl-d-galactoside andN-acetyl-d-mannosamine, for example, in Slomka et al., J ClinPeriodontol. (2017), 44(4):344-352. In one embodiment, the daily dose ofprebiotic is from about 0.00001 g to about 1 g, more preferably fromabout 0.0001 g to about 0.5 g and, even more preferably, from about0.0005 g to about 0.1 g, of the prebiotic.

The term “probiotic” as used herein refers to live, dead, andinactivated microorganisms which, when administered in adequate amounts,confer a beneficial effect on the health or well-being of the host.Examples of such probiotics include substantially pure bacteria (i.e., asingle isolate), or a mixture of desired bacteria. Health benefits mayinclude those relating to cardiovascular health, bone health, guthealth, oral health, skin or dermal health, anti-stress or behavioralhealth, and immune health. For the purpose of the present disclosure,“probiotics” is further intended to include the active metabolitesgenerated by the microorganisms of the present disclosure, if they arenot separately indicated. Such cell metabolites may be obtained by usinglysates of probiotic bacteria or fermentation supernatants. Metabolitesmay include organic and inorganic molecules, alcohols, aldehydes, aminoacids, carbohydrates and components thereof, peptides, proteins andcomponents thereof, extracellular enzymes, cell-wall-bound enzymes,membrane-bound or intracellular enzymes, electron transport moleculesand components thereof, or other cell-wall, membrane or cytoplasmiccomponents and molecules, hormones or hormone like substances, lipids,oils, fats or fatty acids and components thereof, organic acids, nucleicacids or ribonucleic acids and components thereof, carbon compounds,nitrogen compounds, phosphate compounds, pigments, and vitamins as wellas mixtures of any of above components and molecules, for example, inFernandez-Gutierrez et al., (2017) Scientific Reports |7: 11100|DOI:10.1038/s41598-017-11446-z, and for example in MacKenzie et al.,Microbiology (2010), 156, 3368-3378. For the purpose of the presentdisclosure, “probiotics” is further intended to include inactivated ordead probiotic bacteria and yeast such as those used to co-aggregatespecific microorganisms or other prokaryotic or eukaryotic cells andcomponents thereof.

Examples of microorganisms generally recognized as probiotics areAcetobacterium, Acetitomaculum, Bacillus, Bacteroides, Bergeyella,Bifidobacterium, Blautia, Capnocytophaga, Clostridium, Corynebacterium,Enterococcus, Eubacterium, Holophaga, Lactobacillus, Lautropia,Leuconostoc, Moraxella, Moorella, Neisseria, Pasteurellaceae,Prevotella, Ruminococcus, Saccharomyces, Sporomusa, Staphylococcus,Stenotrophononas, Streptococcus, Treponema, Weissella, Wolinella, andXenophilus and mixtures thereof. More particularly, Bifidobacteriumanimalis, Bifidobacterium lactis, Bifidobacterium longum, Lactobacillusbrevis, Lactobacillus helveticus, Lactobacillus johnsonii, Lactobacillusparacasei, Lactobacillus plantarum, Lactobacillus reuteri, Lactobacillusrhamnosus, Lactococcus cremoris or Lactococcus lactis, Saccharomycescerevisiae or Saccharomyces boulardii, Streptococcus salivarius,Streptococcus thermophilus.

Enzymes and proteins may be also used as a health ingredient, such asamyloglucosidase, glucose oxidase or glucosidase, lactoperoxidase,mutanases, dextranases, lipases, laccases, peptidases or proteinases,xylanases, other polysaccharide degrading enzymes, and other hydrolyticenzymes; proteins such as Colostrum (Lactoferrin, sIgA), bacteriocins,lytic phage or components thereof, proteins or other inhibitors ofquorum sensing in target bacteria and other microorganisms. Lipids andderivatives thereof that can be dispensed to a companion animals' mouthsuch as polyunsaturated or omega-3 fatty acids, monounsaturated fattyacids such as 1-tetradecanol complex (e.g. Hasturk et al., 2007, JPeriodontology, Vol 78:924-932), and derivatives of fatty acids, such asthose described in WO 2011/061237. Such publications and/or patentapplications in this specification are incorporated by reference andrelied upon in their entirety.

The term “pharmaceutically acceptable” or “veterinarily acceptable” asused herein refers to molecular entities and compositions that do notgenerally produce an adverse, allergic or other untoward reaction whenadministered to an animal. Moreover, for animal administration, it willbe understood that preparations should meet sterility, pyrogenicity,general safety, and purity standards as required by FDA, USDA, orEuropean Medicine Agency.

The term “palatant,” “palatability enhancer,” “flavoring agents,”“flavoring”, or “flavorants” means any material or substance thatenhances the palatability of a food composition to an animal. A palatantor palatability enhancer can be a single material or a mixture ofmaterials and can be a natural (either unprocessed or processed),synthetic, or part natural and part synthetic material. A palatant canbe added to a composition as an additive that comprises the palatant orthat comprises the palatant together with one or more other functionalor non-functional materials. Palatability enhancers may be made whollyor partially from meat or poultry broth concentrate or spray-driedpowder, hydrolyzed proteins, yeast and/or yeast extract, liver, spices,herbs, sweeteners, or any combination such components.

The terms “treating,” “treatment,” or “to treat” as used herein refersto reversing, alleviating, or inhibiting the progress of a disease,disorder, or condition; or decreasing the probability or incidence ofthe occurrence of a disease, or condition in a subject as compared to anuntreated control population, or as compared to the same subject priorto treatment; or delaying or preventing the symptoms associated with adisease, disorder or condition. As used herein “treating” may also referto preventing the recurrence of a disease, disorder or condition or ofone or more symptoms associated with such disease, disorder, orcondition.

The Delivery System

The delivery systems according to the present disclosure comprise aningestible matrix within which one or more functional ingredient(s) aresubstantially uniformly and completely dispersed and in which solubilityand release characteristics in the delivery of any functionalingredient, is greatly facilitated using whisked egg white foam as adelivery vehicle.

Egg white is a clear viscous dispersion of biologically active proteinsin an aqueous medium surrounding the yolk of an avian egg. The egg whiteof domestic hens' egg is approximately 10% mixed proteins includingovalbumin, ovotransferrin, immunoglobulins, lysozyme, and avidin, amongothers. Egg white can be separated from the yolk by mechanical means andis commonly used in foods as a protein source and as a binder andthickener in processed meats. Egg white can be turned into a foam bywhisking and beating in a manner that causes molecular shear forcing thenormally coiled protein molecules to deform into a linear conformationcreating a tenacious interface with trapped air bubbles. Among otherfood applications egg white foam is used in preparation of a variety ofconfections including marshmallow and nougat in which a boiling sugarsyrup is blended with the whisked egg white, cooking and fixing it, andtrapping the air bubbles creating a light frothy material when cooled.

The process of whisking (or beating) egg white proceeds through severalstages of hardness commonly described as ‘soft,’ ‘firm,’ and ‘stiff’peaks after which if whisking is continued the foam will collapse. Thevolume of whisked egg white is usually about 4 times greater than theoriginal liquid egg white and the trapped air bubbles are usuallymicroscopically dispersed with occasional large visible bubbles. Softand firm peak foams can be blended easily with other solutions or soliddispersions in aqueous medium and in doing so they serve to disperse thetrapped air throughout the combined mass. Stiff peak foams are lesseasily dispersed tending to break into non-homogenous clumps, but theycan be cooked by gently heating after which they form a crisphoneycombed mass of which confectionary meringues are a good example.

While egg white foams are popular and commonly used for culinarypurposes their use in delivery of biologically functional ingredientssuch as nutrients and medicaments is novel, and as disclosed here eggwhite foams facilitate inventive constructs that offer great utilitywith particular classes of functional ingredients designed fortherapeutic and nutritional applications.

As disclosed herein it is possible to formulate a functional ingredientusing whisked egg white foam in a manner such that the functionalingredient is concentrated at the foam-air interface (i.e., on thesurface inside the individual bubbles). Egg white foams with surfaceconcentrated functional ingredients provide a much greater availabledose loading, which can be released much more rapidly as compared to aconventional topical formulation.

Among the various classes of functional ingredients many arewater-soluble, some are only soluble in oil while a few are amphipathicdisplaying partial solubility in oil and water in the manner of asurfactant. The solubility of any functional ingredient greatlyinfluences the appropriate manner of its delivery to a human or animalbody, particularly oral applications where the functional ingredient isintended to affect a response on or in the gastro-intestinal tract andin topical and mucosal applications where the functional ingredient isintended to affect a response on or in the skin or mucosal membranes andby extension on structures emanating from these surfaces such as hair,nails, and teeth.

In general, functional ingredients including medicaments andnutritionals that are freely soluble in water present very littleimpediment to incorporation in any formulation. However, achievingadequate release from formulations is not always as easy becausewater-soluble ingredients will freely disperse throughout the entiretyof the administered dose necessitating absorption or assimilation of theentire carrier to access the functional ingredient. Isolating andconcentrating freely water-soluble ingredients to enhance delivery canbe facilitated using whisked egg white foam wherein the water-solubleingredient is dispersed and fixed at a foam interface with a greatlyexpanded surface area which provides an amplified available dose whenthe foam itself is dispersed in other carriers.

Formulations of poorly soluble functional ingredients normally containexcipients designed to facilitate solubility, absorption and/orresidence time on an intended surface. The available dose of anyfunctional ingredient in such formulations is also limited by the amountof formulation that can be applied to the intended surface and themigration or release of the functional out of the formulation and ontoor into the intended surface. Solubility and release characteristics arecritical parameters in the delivery of any functional ingredient, and asdisclosed herein, both can be greatly facilitated using whisked eggwhite foam as a delivery vehicle.

Oil and oil soluble ingredients and many amphipaths present morechallenging problems in formulation. It is well known in culinary artthat any trace of oil or fat will prevent whisked egg white fromfoaming, equally once egg white has been whisked and foamed, theaddition of trace amounts of fat or oil or oil soluble ingredients willcause the foam to collapse. Egg white foams are normally incompatiblewith any lipophilic (fat soluble) component and this presents atechnical challenge in using such foams for enhanced delivery of oilsoluble or amphipathic functional ingredients.

The use of emulsification techniques to disperse fine droplets ofoil-based ingredients, particularly if such droplets are stabilized byamphipathic excipients, facilitates more flexible incorporation andrelease characteristics from whisked egg white foams.

An emulsion is a dispersion of oil droplets in an aqueous medium (oil inwater) wherein an emulsification agent is used to prevent the oildroplets coalescing. An example of a functional emulsion is provided byFolan in U.S. patent application Ser. No. 15/384,372 based on WO 2011061237 which is fully incorporated herein. In that application waterinsoluble free fatty acids (oils) are emulsified in membrane lipids ofnatural origin, the free fatty acid is preferably Caprylic acid and theemulsification agent is preferably de-lipidised lecithin.

Surface area is a critical a parameter affecting release characteristicsand delivery of any functional ingredient. The example of an ointment onskin serves to illustrate the critical nature of surface area. Thecontact surface of ointment on skin is the delivery interface and theavailable dose of functional ingredient is the amount of the functionalat the contact surface. The available dose is not immediately increasedby adding additional layers of ointment because the functional mustfirst migrate through the additional layers before it is available atthe skin interface. The use of egg white foam as a delivery vehiclegreatly amplifies available surface with consequentially improvedrelease characteristics.

In the context of delivering a functional ingredient it should berecognized that the total surface area of individual foams will varydepending on the extent of whisking and the actual protein concentrationof the starting solution. For the purposes of the disclosure, it issufficient to know that a whisked egg white foam that has approximatelyquadrupled in volume will present a surface area approximately severalhundred times greater than the original protein solution or anyequivalent volume of a non-foamed formulation.

As disclosed herein it is possible to formulate a functional ingredientusing whisked egg white foam in a manner such that the functionalingredient is concentrated at the foam-air interface-that is on thesurface inside the individual bubbles. Egg white foams with surfaceconcentrated functional ingredients provide a much greater availabledose loading, which can be released much more rapidly, compared to aconventional topical formulation.

The delivery systems according to the present disclosure are suitablefor administration to both human and non-human animals. One skilled inthe art will appreciate that each delivery system can be formulateddifferently according to the type of animal to which it is to beadministered. For example, for administration to an animal such as a cator a dog, meat or fish-based flavors and flavorants may be added. Foradministration to a human, the delivery system may be formulated, forexample, as a confectionery using fruit-based or other flavors. Thedelivery systems are especially suited for oral administration due totheir palatability. Additionally, due to the highly portable format, thedelivery systems are simple and convenient to administer and to consumefor both humans and other animals.

The delivery systems of the present disclosure can be tailored forspecific purposes; thus, the delivery systems can be formulated withspecific combinations of functional ingredients in order to producespecific physiological effects. For example, a drug delivery system canbe formulated to contain certain combinations of drugs or diagnosticagents. Other delivery systems can be formulated with combinations offunctional ingredients for example to promote endurance, promotecardiovascular health, control fat and/or cholesterol, promote healthyjoints, mainain or improve bone density, enhance cellular antioxidantcapacity, or control appetite.

The delivery systems of the present disclosure comprise one or morefunctional ingredient(s) substantially uniformly dispersed within amatrix which generally comprises 1) egg white at a protein concentrationthat forms a foaming matrix when whipped; 2) one or more heat resistantand/or heat sensitive gelling agents; 3) a pH regulator; 4) one or moreplasticizers and/or humectants; 5) one or more sources of water.Moisture content and controlling the physical characteristics of theresidual moisture within the matrix is facilitated by inclusion of oneor more gelling agents. Additives such as natural or artificialflavorings, colorings, acidulants, buffers, and sweeteners can beincluded in conventional amounts in the matrix.

The delivery systems of the present disclosure can be formulated suchthat the matrix has a final pH in the range of about 2.5 to about 8.5.In one embodiment, the matrix has a final pH of between about 3.0 andabout 8.5. Acidic pH is known in the art to promote degradation ofcertain functional ingredients. For delivery systems formulated todeliver functional ingredients which are sensitive to, or reactive at,acidic pH, therefore, the final pH of the matrix is neutral to mildlybasic. By neutral to mildly basic pH it is meant that the final pH isbetween about 6.0 and about 8.5. For those functional ingredients thatare more stable in acidic form, such as trimethylglycine, or functionalingredients which may react with other components at neutral pH such asglucosamine hydrochloride, the pH of the matrix of the delivery systemsmay have a final pH below neutral.

In one embodiment of the present disclosure, the delivery systems areformulated such that the matrix has a final pH of about 5 and thus aresuitable for delivery of functional ingredients that are stable and/orunreactive at acidic pH.

As described in the Examples below, formulations containing foamed eggwhite will typically contain other water-soluble constituents such asgels, polymers, and organic acids which when cooked together with theegg white protein constitute the solid scaffold structure of the foamwhich is infused with air bubbles. When oil-based ingredients, or oil inwater emulsions are added to the solid matrix there is phase repulsionbetween the aqueous medium and the oil droplets causing these dropletsto migrate to a position of least resistance which is the water-airinterface inside the trapped bubbles.

Because of the greatly expanded surface area and the concentration ofoil droplets at the surfaces of the foam bubbles, formulationsincorporating emulsified oils will have much greater available doseloading and much faster release characteristics compared to a similarmass of non-foamed formulation.

The texture, physical attributes, form, and shape of the matrix asdescribed below, can be varied by altering the ratio of ingredientswithin the given ranges using the methods described herein or by methodsfamiliar to a worker skilled in the art. In addition, the specificselections of the possible components provided below, must be safe foranimal and/or human consumption and meet regulatory standards, such asthose of the Codex Alimentarius.

Egg white foam formulations intended to deliver a functionally activeemulsion have great utility in topical skin and mucosal healthcare whererapid release of available dose loading is desirable because of shorttransit times on the mucosa of the mouth for example. Exemplaryemulsions comprise (a) one or more saturated or unsaturated free fattyacids having from 4 to 22 carbon atoms or a pharmaceutically acceptablesalt thereof; and (b one or more delipidised membrane lipids, asemulsifying agent for the free fatty acid(s) or the salt thereof.

Modified formulations of the same type may be used to protect emulsiondroplets using whisked egg white as a protein coat to delay gastricdigestion and achieve enhanced intestinal availability and absorption offunctional ingredients carried in the emulsified oil. Protein digestionin the mammalian stomach is largely due to pepsin activity at low pH.Some of the individual proteins in egg white particularly ovalbumin andovomucoid are particularly resistant to pepsin digestion, the additionof lecithin to egg white adds to pepsin resistance and whisking prior tocooking facilitates intact gastric transfer. Once in the duodenumtrypsin proteolysis will affect rapid release of the encapsulated oilemulsion and the availability of these for absorption at theenterocytes.

Emulsions of the type described above are not confined to the use offree fatty acids. Emulsification techniques may be used to deliver oilsoluble ingredient either on their own or in solution in other oils suchas neutral triglycerides of which Miglyol 812N from IOI Oleo, Hamburg,Germany, is one example. The same oil-water phase repulsion forces willapply to concentrate the oil droplets at the water-air interface insidethe bubble, regardless of the type of oil in the emulsified droplet andregardless of any oil soluble ingredient in the oil. Examples of otheroil soluble functional ingredients that would benefit from enhanceddelivery in emulsions dispersed in whisked egg white foam include butare not limited to antibiotics like Mupirocin, antimycotics likeClotrimazole, antiseptics like Chlorohexidine, anti-inflammatories likeKetoprofen and nutritionals like the oil soluble vitamins A, D, E and K.

Whisked egg white foam is especially useful for incorporation anddelivery of functional ingredients that exhibit amphipathic properties,these include local anesthetics such as Lidocaine, Biocides such asBenzalkonium Chloride, antimicrobial agents such as Delmopinol andanti-inflammatory agents such as curcumin. When dispersed in egg whitebefore or after foaming, the hydrophilic aspect of an amphipathicmolecule will tend to associate with the water based proteinaceousmatrix while the lipophilic aspect (also hydrophobic) will naturallyorientate at the water-air interface and/or with the lipophilic aspectof adjacent molecules encouraging the establishment of micellar andlamellar structures all of which can be used to engineer improvedrelease characteristics.

In embodiments, lecithin is used in combination with whisked egg whitefoam. Lecithin is an amphipathic molecule which may be extracted fromvegetable sources like soy and from egg yolk where it is closelyassociated with but distinctly separate from the egg white. Asdemonstrated herein lecithin may be combined with egg white beforewhisking where despite its amphipathic nature it has very little impacton the foaming properties of the egg white but from where it greatlyfacilitates inclusion and release of other amphipathic and lipophilicagents as detailed previously including but not limited to Delmopinol,curcumin, Lidocaine and Bezalkonium Chloride.

In other embodiments, lecithin is used in combination with whisked eggwhite to facilitate the construction of free fatty acid emulsions insitu as part of the formulating process While foamed egg white isincompatible with oil and will collapse if even a trace amount is addeda method whereby specific oils such as free fatty acids in the form oftheir water-soluble sodium or potassium salts may be combined with theegg white before whisking without significantly affecting the foamingproperties has been developed utilizing a suitable amount of anemulsifying agent like de-lipidised lecithin being added to the eggwhite. The salts of free fatty acids may be converted back to theirprotonated oil form by acidification after the foam has been integratedinto the finished formulation, without affecting the foamedcharacteristics.

De-lipidised lecithin is amphipathic and exerts superior emulsifyingproperties by virtue of one facet being oil soluble and an opposingfacet of the same molecule being water-soluble. De-lipidised lecithinmay be added to an egg white protein solution and if allowed a suitabletime to hydrate it disperses homogenously throughout.

In embodiments, a water-soluble salt of Caprylic acid such as sodiumCaprylate (sodium octanoate) may also be added in an appropriate ratiowith the de-lipidised lecithin. The egg white protein/de-lipidisedlecithin/sodium Caprylate mix may be whisked, the texture of the blendedfoam is more viscous because of the added ingredients but the extent ofair incorporation is approximately the same. Generally, the blended eggwhite foam is added to and cooked in a gel formulation after which anamount of an organic acid such as citric or ascorbic acid is added tothe formulation in a molar equivalent amount relative to the amount ofsodium Caprylate. The effect of the acidification is to drop the pHbelow the dissociation constant (pKa) of sodium Caprylate at which pointthe salt is converted to its water insoluble free fatty acid oil. SodiumCaprylate is intimately dispersed with de-lipidised lecithin in the eggwhite before being whisked and it remains intimately dispersed duringand after whisking and during incorporation and cooking of the foam inthe gel formulation. When the formulation is acidified below theappropriate pH value the intimately dispersed sodium Caprylate isconverted to its free acid oil and associates by phase attraction withthe lipophilic facet of the co-dispersed lecithin in a form similar toan emulsified oil droplet which is located at or at least very close tothe air interface of the whisked egg white bubble.

It will be appreciated that any functional ingredient that does notinterfere with the whisking and foaming of egg white may be incorporateddirectly during the whisking process. And further, incorporation at thefoam air interface will enhance delivery of any compatible ingredientdue to amplified available dose and improved release characteristics atthe expanded surface area of the foam.

The foamed egg white matrix can optionally contain other additives suchas sweeteners, chelating agents, flavorings, colorings, modifiedvegetable gums or celluloses, or a combination thereof that does notinterfere with the whisking and foaming of egg white. It will be readilyapparent that additives for inclusion in the matrix should be selectedsuch that they do not affect the properties of the matrix, do notexhibit substantial reactivity with the functional ingredients in thematrix, and are stable during preparation of the matrix.

The sweetener can be selected from a wide variety of suitable materialsknown in the art. Representative, but non-limiting, examples ofsweeteners include sugars including but not limited to sucrose,fructose, lactose, sorbose and glucose and alcohol derivatives of sugarsincluding but not limited to glycerol, xylitol, sorbitol, lactitol anderythritol as plasticizers with or without additional gelling agentslend further utility in construction of whisked egg white formulations.The ratio of egg white to selected sugars and/or sugar derivatives maybe in the range of 1.0:0.1 to 2.0:10.0 or from 1.0:5.0 to 1:1.

Metallic salts and free metal ions such as magnesium, calcium, zinc, andiron are frequently problematic and may inhibit reactive functionalingredients such as free fatty acids. To counteract the effect of saltsand free ions, a chelating agent may be added to the egg white prior towhisking and or to other constituents in the gel scaffold. Suitablechelating agents include but are not limited to ortho-phosphates andpolyphosphates such as di-sodium or di-potassium orthophosphate,di-sodium uridine monophosphate, sodium phytate and sodiumhexametaphosphate. Non-phosphate based chelating agents includetri-sodium citrate and Ethylenediaminetetraacetic acid. Incorporation ofa chelating agent may be from 0.1% W/V to 5.0% W/V.

Some chelating agents inhibit microbial growth due to sequestration ofessential mineral metabolites particularly in environments where theseare available in very low concentration. Usually, microbial growth isrestored when mineral supply is supplemented although it wasunexpectedly discovered, as exemplified herein, that in particularembodiments the inclusion of other functional ingredients such asemulsions of free fatty acids will act synergistically with a chelatingagent to suppress microbial growth in the presence of excess mineralsupplementation.

It will be appreciated by those skilled in the art that certainphysiological environments have characteristically higher mineralconcentrations, these include blood, serum, mucus, and saliva wheredi-valent metal ions are essential. In the treatment of wounds forexample it may be desirable to reduce the concentration of di-valentions such as calcium to inhibit blood clotting and in the event that amicrobicidal effect was also desirable a combination of a chelatingagent such as hexametaphosphate with a functional emulsion of free fattyacid would lend great utility.

Equally it is well known that saliva is super saturated with respect tocalcium ion, and furthermore that non-selective deposition of calcium indental plaque results in tenacious accretions of calculus which greatlyexacerbate the risk of gum disease. In formulations that are intended topromote oral health, and or treat or prevent disease of the oral cavity,it may be an advantageous to use a calcium chelating agent to reducecalculus. Where it is desirable to include an antimicrobial effect tolimit dental plaque formation the synergistic combination ofhexametaphosphate and a functional emulsion of free fatty acid achievessignificantly enhanced health benefits.

Chelating agents such as polyphosphates are commonly used in skin careformulations to stabilize and prevent degradation reactions commonlycatalyzed by di-valent metal ions. In medicated cosmetic applicationsfor example where it is desirable to achieve an additional antimicrobialeffect a synergistic combination of a polyphosphate and a functionalfree fatty acid emulsion is particularly beneficial. One example of suchmedicated cosmetics is a skin cream for acne and a further example is ashampoo designed to ameliorate infectious dandruff.

Suitable flavorings that can be added to the delivery system include,both synthetic flavor oils and oils derived from various sources, suchas plants, leaves, flowers, fruits, nuts, and the like. Representativeflavor oils include spearmint oil, peppermint oil, cinnamon oil, and oilof wintergreen (methylsalicylate). Other useful oils include, forexample, artificial, natural, or synthetic fruit flavors such as citrusoils including lemon, orange, grape, lime, and grapefruit, and fruitessences including apple, strawberry, cherry, pineapple, banana,raspberry, and combinations thereof.

The amount of flavoring agent employed is normally a matter ofpreference subject to such factors as concentration/dilution of theflavor stock, flavor type, base type, and strength desired. In general,amounts of about 0.01% to about 50% weight of a final product areuseful.

In one embodiment of the present disclosure, vanillin is included in thematrix as a flavoring agent in amounts of about 1.5% In anotherembodiment, the flavoring agent is added in amounts of about 0.03% toabout 15%.

Colorings suitable for use in foodstuffs can be optionally included inthe matrix to add aesthetic appeal. A wide variety of suitable foodcolorings are available commercially, for example, from Warner Jenkins,St. Louis, Mo. Where a synthetic coloring agent is used in the matrix,the amount ranges from about 0.01% to about 2% by weight.

In one embodiment of the present disclosure, a synthetic coloring agentis added to the matrix in an amount between about 0.03% to about 1% byweight.

Due to the substantially uniform and complete dispersion of thefunctional ingredients within the matrix, the delivery systems aresuitable for division into sub-units. For example, if a single unit of adelivery system of the disclosure is divided into three subunits, eachsubunit will contain a third of the dose of the original unit. Suchdivision would not be possible with other delivery systems in which thefunctional ingredients are not evenly dispersed.

In embodiments the foamed egg white matrix is used as part of a dualaction dental chew for dogs. This dual-action chew works like atoothbrush and toothpaste to keep dog mouths clean and protected fromharmful bacteria. The dental chew comprises two components; a flexiblebase and a filling. The flexible base is shaped and designed to reduceplaque and calculus through mechanical action (scrubbing, abrasion)during the chewing process. In addition, the chew base features a‘reservoir’/cavity for the filling that contains the functionalingredient(s). The filling comprises the foamed egg white matrix and atleast one functional ingredient, and wherein the filling and thefunctional ingredient is distributed in the mouth during the chewingprocess, which provides a mechanical barrier, preventing bacteria fromadhering to surfaces in the oral cavity (teeth, tongue, and gums).

In embodiments the dental chew base is bone shaped and has ridges andnodules on at least a part of the exposed surface area to enhancemechanical cleaning.

In embodiments the functional ingredient in the filling comprises anemulsion comprising (a) one or more saturated or unsaturated free fattyacids having from 4 to 22 carbon atoms or a pharmaceutically acceptablesalt thereof; and (b) one or more delipidised membrane lipids, asemulsifying agent for the free fatty acid(s) or the salt thereof

Materials and Methods.

The construction of a whisked egg white foam with or withoutincorporation of functional ingredients such as salts of free fattyacids and/or lecithin is based on an aqueous dispersion of egg whiteproteins.

The use of fresh egg white is not particularly suitable for industrialscale process and commercially available powdered egg white offers muchgreater convenience as well as an opportunity to vary the concentrationof protein in the foam. Egg white powder is available from many sourcesincluding Canadian Inovatech, Abbotsford, BC, Canada.

Egg white powder is re-hydrated in water that has been purified byreverse osmosis. A 10% W/W dispersion of egg white protein is 10 gramsof powdered egg white in 90 grams of water. Depending on the type offoam required and the amount of other ingredients that are incorporated,dispersions of up to 40% protein may be re-hydrated. Higher proteinconcentrations do not build as much volume during whisking but lendthemselves to other methods of foaming including the use of hydrogenperoxide and catalase enzyme.

Whisked and foamed egg white forms a protein scaffold incorporating asignificant amount of trapped air which needs to be fixed by heating toabout 80° C. to denature and render the scaffold insoluble. Heattransfer to the protein scaffold is facilitated by moisture content andcontrolling the physical characteristics of the residual moisture isfacilitated by inclusion of other gelling agents (hydrocolloids).Hydrocolloids are hydrophilic polymers of vegetable, animal, microbialor synthetic origin naturally present or added to aqueous foodstuffs fora variety of reasons due to their unique textural, structural andfunctional properties. In general, they are used for their thickening,gelling properties, and/or heat resistance as well as their waterbinding and organoleptic properties. Hydrocolloids can also be used toimprove and/or stabilize the texture of a food product while inhibitingcrystallization. Examples of hydrocolloids include but not limited tostarch, tragacanth, gluten, fumed silica, polyethylene glycol, celluloseand cellulose derivatives, gelatin, collagen, mucins, pectin, gumarabica, guar gum, acacia gum, karaya gum, locust bean gum, xanthan gum,carrageenan, agar, gellan and/or sodium alginate and combinations ofthese.

The selection of the hydrocolloid to be used in the matrix will dependon the pH of the matrix and the texture and consistency required for thefinal product. The type of hydrocolloid used will also affect the settemperature of the matrix. For example, the use of a gelatin/gellanmixture or a gelatin/pectin mixture provides a set temperature around35° C., whereas the use of carrageenan or locust bean gum will result ina set temperature closer to 60° C., and whereas the use of agar willresult in a set temperature closer to 45° C., Thus, the choice ofhydrocolloid for use in the matrix is also dependent upon the propertiesof the functional ingredient(s) to be incorporated into the deliverysystem. Functional ingredients that are unstable at higher temperatureswill require the selection of a hydrocolloid or mixture of hydrocolloidsthat have a low set temperature, whereas functional ingredients that aremore stable can be used with hydrocolloids having a higher settemperature.

In one embodiment of the present disclosure, the matrix comprisesgelatin. The term “gelatin” refers to a heterogeneous mixture ofwater-soluble proteins of high average molecular weight derived from thecollagen-containing parts of animals, such as skin, bone and ossein byhydrolytic action, usually either acid hydrolysis or alkalinehydrolysis. Different types of gelatin can be prepared by altering theprocess parameters. Gelatin is defined generally using a “Bloom value”which indicates the strength of the gel formed under certaincircumstances using the gelatin. In the preparation of confectionery,when a harder gel is desired, gelatin having a higher Bloom value isused. Conversely, when the final product is required to be more flowing,gelatin having a lower Bloom value is used. The water holding capacityof gelatin alone is lower than that of a combination of gelatin withanother hydrocolloid, such as gellan or pectin, and may necessitate theuse of a higher amount of gelatin to achieve the desiredgelation/texture of the matrix. When the hydrocolloid in the matrix ofthe present disclosure comprises gelatin, the Bloom value (BL) isgenerally about 100 to 300BL.

In one embodiment, the Bloom value is about 260 BL. In anotherembodiment, a mixture of gelatins with different Bloom values is used.

As indicated above, gelatin can be combined with one or more otherhydrocolloid(s) to impart slightly different characteristics to thematrix. For example, combinations of gelatin with agar, gelatin withpectin, or gelatin with agar and pectin provide a good texture to thematrix. Other combinations of hydrocolloid(s) are also contemplated, forexample but not limited to, agar combined with pectin. When combinationsof gelatin and agar are used in the preparation of the matrix, the ratioof gelatin:agar is typically in the range between about 1:1 to about10:1. These relative amounts provide a cohesive structure to thedelivery system.

In one embodiment of the present disclosure, a combination of gelatinand agar is used in the preparation of the matrix in a gelatin:agarratio of about 1:1 to about 3:1.

In embodiments, the total amount of hydrocolloid incorporated into thematrix is generally between about 0.1% and about 7.0% by weight. In oneembodiment, the total amount of hydrocolloid in the matrix is betweenabout 0.5% and about 6.8% by weight. In another embodiment, the totalamount is between about 1.0% and about 6.0%. In other embodiments, it isbetween about 2.0% and about 6.0%, between about 4.0% and about 6.0%,between about 5.0% and about 6.0% and between about 6.0% and about 7.0%.

In embodiments, the ratio of whisked egg white to selected gellingagents may in the range of 2:1 to 0.2:10, or from 0.1:1 to 1:1.

In other embodiments, the ratio of egg white to gelling agents may rangefrom 0.01:10 to 1.0:10 or from 1.0:10 to 10:1.

A suitable grade of gelatin is 260 bloom 40 mesh available from PBLeiner, Belgium and Food grade Agar and other gelling agents areavailable from many sources including Special Ingredients Ltd,Chesterfield, UK.

Consideration must be given to the impact of the gelling agents onrelease characteristics and on the physical stability of the finishedformulation in practical applications.

In embodiments, other ingredients used in formulations incorporatingwhisked egg white matrix include Caprylic acid or sodium Caprylate whichare available from Merck Chemicals.

In embodiments, the ratio of Caprylic acid to egg white may range from0.01:10.0 to 1.0:10 or from 0.1:1.0 to 1.0:1.0 In embodiments, otheringredients used in formulations incorporating whisked egg white matrixinclude lecithin.

A suitable grade of purified lecithin is available from Lipoid A G,Zurich, Switzerland. In embodiments, the ratio of lecithin to egg whitemay range from 0.01:10.0 to 1.0:10 or from 0.1:1.0 to 1.0:1.0.

Assay of Functional Efficacy

It should be appreciated that the disclosures herein relate to deliveryof a wide range of functional ingredients providing enhanced utility inhuman and animal healthcare including but not limited to therapeutics,prophylactics, and nutritionals.

To illustrate the utility of formulations based on whisked egg white thefollowing Examples make use of an antimicrobial emulsion as disclosed inFolan U.S. patent application Ser. No. 15/384,372. This functionalingredient exerts a dual antimicrobial effect by limiting adhesion ofmicrobial species and reducing their viability through a secondarymicrobicidal/microbistatic effect. For comparison purposes the assay ofmicrobicidal/microbistatic effect is used here and the method isdescribed below.

The assay is a standard microbiological suspension test wherein knownconcentrations of late log phase bacteria, yeast, or fungi areinoculated into a fixed volume or weight of a test substance, blank, orcontrol. After a set period a neutralizing solution is added to stop theantimicrobial effect and the residual population of viablemicroorganisms is enumerated by serial dilution and plate counting. Thecounting procedure is a standard and basic microbiological procedure forenumerating viable microorganisms and will be well known to thoseskilled in the art.

In its generic form the method requires inoculation of 1 gram or 1 ml oftest sample with 0.1 ml of 18 hour (late log phase) bacterial culturefollowed by vigorous agitation to mix. After the pre-determined exposuretime has elapsed, 9.0 ml of neutralizing buffer is added and mixed. Thishas the effect of stopping the microbicidal effect which allows reliableestimates to be made of the percentage kill achieved by a test sample inthe period between inoculation and neutralization. Typically, exposureperiods will range from 30 seconds up to 30 minutes and may progress toseveral hours where that time period is required to measure the effect.In order to enumerate residual viable cells and from that to computepercentage kill, the number of viable cells in the inoculum isdetermined by serial dilution and plate counting. Appropriate blanks andcontrols are used to ensure validity of the neutralizing procedure andto allow for any interference from other constituents in the testsample.

In the assays described herein, the test organism is a standardindicator bacterium, Staphylococcus aureus NCTC 8325-4 (NationalCollection of Type Cultures, Public Health England, Porton Down,Salisbury, UK) known for its tenacious biofilm forming capability.Stocks of bacteria are routinely stored on beads in 50% glycerol at −80°C. When required for viability/microbicidal assay, small aliquots fromthese stocks are spread on an appropriate nutrient agar, grown andsub-cultured to ensure purity. Where broth cultures are required, 250 mlErlenmeyer flasks containing 100 ml of broth are inoculated with atransfer loop from pure agar cultures and incubated under constantagitation in a rotary incubator at 37° C.

Indicator bacteria are routinely cultured using brain heart infusion(BHI) broth and agar or tryptone soya agar or broth (TSB), both of whichmay be acquired commercially from Oxoid, UK. The diluting andneutralizing buffers used in the present method are phosphate bufferedsaline (PBS), containing 137 mM sodium chloride, 2.7 mM potassiumchloride and 10 mM phosphate, to which is added 3% polysorbate Tween 80(anionic surfactant), 0.3% lecithin, and 0.5% histidine as neutralizingagents. These ‘neutralizing’ agents are those prescribed under EUGuidelines for ISO certification of microbicidal efficacy and werevalidated as suitable for neutralizing free fatty acids at theconcentrations used herein.

Test samples as prepared in the following Examples are assayed by firstdissolving or dispersing a measured quantity of sample in a measuredamount of sterile water. Typically, one gram of sample will be maceratedin 1 gram of water which represents a 50% dilution of the sample and itseffective dose load. The macerated sample is inoculated with 1 ml of18-hour (late log phase) culture of the indicator organism, stirred andincubated at 37° C. for fixed periods of time after which 9 ml ofneutralizing buffer is added and mixed by inversion. The number ofviable cells in the test sample after incubation and neutralization ismeasured by serial dilution and plate counting and compared to thenumber of viable cells in 1.0 ml of control inoculum treated in exactlythe same way with test sample or with a blank test sample containing nofunctional ingredient.

Typically an overnight culture of the indicator organism will contain inexcess of 8 logs of viable cells per ml (1.0×10⁸ or 100,000,000) per ml.Typically a 2% W/W dose loading of the free fatty acid emulsion used inthe following Examples will achieve greater than 90% or 1 log reductionin viability in 30 seconds of exposure and it is not unusual to seegreater than 6 logs (99.9999%) reduction in 5 minutes.

EXAMPLES

These delivery systems and methods of making the delivery systems areexemplary only, other formulations and methods will be apparent to oneof ordinary skill in the art, and such other formulations and methodsare intended to be included within the scope of this invention.

Example 1—Egg White/Gelatin/Sorbitol Formulation Using an Oil in WaterEmulsion as a Functional Ingredient

TABLE 1 Foamed egg white matrix formulation of Example 1. IngredientWeight in grams 1 Purified water 75 2 Citric Acid 0.5 3 Egg white at 10%protein 120 concentration 4 Gelatin 260 Bloom 16 5 Sorbitol 60 6Powdered flavoring 4 7 Functional emulsion based on 4 U.S. patentapplication 15/384,372 Total weight 279.5

The foamed egg white matrix comprising a functional emulsion was madeaccording to the following process;

-   -   (a) Ingredients 1 (purified water), 2 (Citric acid), 3 (egg        white), and 4 (gelatin) from Table 1 were combined in the        amounts delineated in a suitable vessel at room temperature and        allowed about 30 minutes to fully hydrate the gelatin,    -   (b) using suitable scale equipment with whisk attachments the        mass was carefully beaten/whisked for approximately 5 minutes to        achieve a foam mass that holds its shape when pulled        upwards—firm peaks,    -   (c) the mass was heated (using a water-bath or Bain Marie) while        stirring constantly and using a suitable thermocouple in the        foam and bringing the mass up to about 80° C.,    -   (d) ingredient 5 (Sorbitol) was added in the amount delineated        in Table 1 and stirred in. The foam mass cools to approximately        60° C. due to the endothermic solubilization of the sorbitol,    -   (e) the temperature was set/held at 60° C. (using for example a        water bath/Bain Marie) and maintained the foam mass at this        temperature,    -   (f) ingredient 6 (flavoring) was added in the amount delineated        in Table 1 and stirred in, and    -   (g) ingredient 7 (functional emulsion) was added in the amount        delineated in Table 1 and stirred in.

The foam mass can be held at 60° C. for periods of up to 5 hours withoutsignificant deterioration. The foam mass can be dispensed to molds andallowed to cool where it forms a firm flexible material.

Assay of Antimicrobial Effect:

The total weight of the formulation in Example 1 was 279.5 gramscontaining 4 grams of functional emulsion (1.4%). The functionalemulsion contained 10% free Caprylic acid in the oil phase and so theconcentration of free Caprylic acid in the formulation of Example 1 was0.14%.

The Antimicrobial assay as described in the Materials and Methodssection above gave the following result:

Inoculum of Staphylococcus aureus  1.7 × 10⁹ Viable cells per ml, Zerotime in the assay 1.35 × 10⁶ Viable cells per ml, One Minute Exposure 2.2 × 10⁴ Viable cells per ml, and One Minute Log Reduction 1.15 Logs.

Example 2—Egg White/Gelatin/Agar Formulation Using an Oil in WaterEmulsion as a Functional Ingredient

In this example a combination of gelatin and agar was used to modulatethe solubility and thermal tolerance of the whisked egg whiteformulation. Gelatin starts to melt at temperatures in the region of 35°C. and the stability of the finished formulation having only gelatin asa gelling agent is subject to flow and deformation if storagetemperatures exceed the melting temperature of gelatin (35° C.). Agar isa polysaccharide gel which melts at 80° C. and remains liquid attemperatures down to 45° C. In combination with gelatin thecharacteristic hysteresis of agar can be used to improve thermalstability of formulations without losing the low temperaturesolubilization of the gelatin. Glycerol is added in this Example toprevent excessive drying, its humectant effect retains residual water.

TABLE 2 Foamed egg white matrix formulation of Example 2. IngredientWeight in grams 1 Purified water 300 2 Citric Acid 1.0 3 Agar 7.6 4Gelatin 260 bloom 20 5 Purified Water 50 6 glycerol 25 7 Egg white at20% protein 50 concentration whisked to firm peaks 8 Flavoring powder 89 Functional emulsion based on 10 U.S. patent application 15/384,372Total weight 471.6

The foamed egg white matrix comprising a functional emulsion was madeaccording to the following process;

-   -   (a) Ingredients 1 (purified water), 2 (Citric acid), 3 (agar),        and 4 (gelatin) from Table 2 were combined in the amounts        delineated in a suitable vessel and allowed about 30 minutes to        fully hydrate then the mixture was heated to about 90° C. using        a thermocouple in the mass to confirm temperature,    -   (b) in a separate vessel—ingredients 5 (purified water) and 6        (glycerol) from Table 2 were combined in the amounts delineated        in a suitable vessel,    -   (c) using suitable scale equipment with whisk attachments        carefully ingredient 7 (20% egg white) from Table 2 in the        amounts delineated was whisked/beaten in a suitable vessel for        approximately 5 minutes to achieve a foam mass that holds its        shape when pulled upwards—firm peaks.    -   (d) When the mixture from step (a) reached 90° C. and the agar        had melted the glycerol-water mixture from step (b) was added        and stirred well to mix. The temperature will drop to        approximately 70° C., and step (e) was immediately performed,    -   (e) 50 grams of the mixture from step (c) was added to the        mixture from step (d) and stirred in vigorously, and        continuously stirred while heating until temperature exceeded        80° C. upon which heating was turned off and/or the mixture was        removed from heating,    -   (f) ingredient 8 (flavoring) from Table 2 was added in the        amounts delineated and stirred in, and    -   (g) ingredient 9 (functional emulsion) from Table 2 was added in        the amounts delineated and stirred in.

The foam mass can be held at 60° C. for periods of up to 5 hours withoutsignificant deterioration. The foam mass can be dispensed to molds andallowed to cool where it forms a firm flexible material.

Assay of Antimicrobial Effect

The total weight of material in formulation in Example 2 was 471.6 gramscontaining 10 grams of functional emulsion (2.12%). The concentration ofCaprylic acid in the functional emulsion was 10% and so theconcentration of free Caprylic acid in the formulation was 0.17%.

The Antimicrobial assay as described in the Materials and Methodssection above gave the following result:

Inoculum of Staphylococcus aureus 2.8 × 10⁹ Viable cells per ml, Zerotime in the assay 1.9 × 10⁶ Viable cells per ml, One Minute Exposure 2.8× 10⁴ Viable cells per ml, and One Minute Log Reduction 1.61 Logs.

Example 3—Construction of a Formulation with Functional EmulsionPrecursors in the Egg White Foam

In this example a combination a whisked egg white foam containinglecithin and sodium Caprylate as precursors of the functional emulsiondisclosed in Folan U.S. patent application Ser. No. 15/384,372 wasconstructed. The precursors were converted to the active form of theemulsion after the cooking stage using an organic acid to convert theinactive water-soluble sodium Caprylate to the active oil solubleCaprylic acid. Apart from dispensing with the need to separatelyconstruct the functional emulsion, building precursors into the eggwhite foam before it is cooked gives much greater assurance that thefunctional ingredients are concentrated in the foam air interface aftercooling. Separately adding the fully formed emulsion depends onmigration driven by phase repulsion to concentrate the oil droplets atthe surface of the foam bubbles.

TABLE 3 Foamed egg white matrix formulation of Example 3. IngredientWeight in grams Egg white precursor 1 Water 88.28 2 Lecithin S75 fromLipoid AG 0.24 3 Sodium Caprylate from Merck 1.48 4 Egg white powder 10Gel base 5 Purified water 300 6 Agar 9 7 Gelatin 260 bloom 10 8 PurifiedWater 50 9 glycerol 25 10 Egg white precursor whisked 50 to firm peaks11 Flavoring powder 8 12 Citric acid 2.2 Total weight 454.2

The foamed egg white matrix comprising precursors to a functionalemulsion was made according to the following process;

-   -   (a) Ingredients 1 (Water) and 2 (lecithin) from Table 3 were        combined in the amounts delineated in a suitable vessel at room        temperature and allowed about 10 minutes to hydrate,    -   (b) ingredient 3 (sodium Caprylate) from Table 3 was added in        the amounts delineated and allowed sufficient time for full        solubilization before proceeding,    -   (c) ingredient 4 (egg white powder) from Table 3 was added in        the amounts delineated and wetted fully using a spatula, and        allowed about 30 minutes to fully hydrate,    -   (d) in a separate suitable vessel ingredients 5 (purified        water), 6 (agar), and 7 (gelatin) from Table 3 were combined in        the amounts delineated and allowed about 30 minutes to fully        hydrate then the mixture was heated to about 90° C. using a        thermocouple in the mass to confirm temperature,    -   (e) while the mixture from step (d) was coming up to temperature        using suitable scale equipment with whisk attachment the fully        hydrated composition from step (c) was whisked/beaten for        approximately 5 minutes to achieve a foam mass that holds its        shape when pulled upwards—firm peaks forming ingredient 10,    -   (f) when the mixture from step (d) reached about 90° C. and the        agar had melted ingredients 8 (purified water) and 9 (glycerol)        from Table 3 were combined in the amounts delineated in a        separate vessel and added to mixture from step (d), stirred        well. The temperature will drop to approximately 70° C. and        step (g) was immediately performed,    -   (g) 50 grams of the mixture from step (e)(ingredient 10) was        added to the mixture from step (d) and stirred in vigorously        while heating, and continuously stirred until the temperature        exceeded 80° C. upon which heating was turned off and/or the        mixture was removed from heating,    -   (h) ingredient 11 (flavoring) from Table 3 was added in the        amounts delineated and stirred to dissolve, and    -   (i) ingredient 12 (citric acid powder) from Table 3 was added in        the amounts delineated and stirred well and allowed sufficient        time to dissolve and the pH was checked to ensure it is less        than 5.0.

The foam mass can be held at 60° C. for periods of up to 5 hours withoutsignificant deterioration. The foam mass can be dispensed to molds andallowed to cool where it forms a firm flexible material.

Assay of Antimicrobial Effect:

The total weight of the formulation in Examples 3 was 454.2 gramscontaining 0.736 grams of sodium Caprylate (0.16%) in 50 grams ofwhisked egg white. The molecular weight of sodium Caprylate was 166.19and that of Caprylic acid was 144.21, the conversion factor 1.15 and soif all the dose load of sodium Caprylate was converted to free Caprylicacid on acidification the concentration of free Caprylic acid in theformulation was 0.14%.

The Antimicrobial assay as described in the Materials and Methodssection above gave the following result:

Inoculum of Staphylococcus aureus 1.83 × 10⁹ Viable cells per ml, Zerotime in the assay 2.54 × 10⁶ Viable cells per ml, One Minute Exposure1.97 × 10³ Viable cells per ml, and One Minute Log Reduction 2.53 Logs.

Example 4—Alternative Method of Developing an Egg White Foam

In some situations where suitable whisking equipment is not readilyavailable it is possible to develop an adequate protein foam usinghydrogen peroxide and a catalase enzyme. A much more concentrated eggwhite protein is required together with gelatin which has been fullyhydrated at room temperature. Hydrogen peroxide is available as 8%, 16%and 32% solutions, the higher concentrations presenting a significantchemical burn hazard if accidentally splashed on human skin. The optimalamount of hydrogen peroxide depends on the concentration in solution andthe ambient temperature. Catalase enzymes are commercially available andapproved for food use. A very small amount of the enzyme is required toactivate decomposition of the peroxide generating a relatively largeamount of oxygen which is dispersed throughout the egg white gelatinmass. Continuous stirring is required to maintain the foam and heatingto cook should be started as soon as the peroxide reaction has stopped.

TABLE 4 Foamed egg white matrix formulation of Example 4. IngredientWeight in grams 1 Purified water 75 2 Gelatin 260 Bloom 10 3 Egg whiteat 40% protein concentration and water 100 4 16% hydrogen peroxide 10 5Catalase 929L from Biocatalysts, Cardiff, UK 0.2 6 Sorbitol 60 7Powdered flavoring 4 8 Functional emulsion based on 4 U.S. patentapplication 15/384,372 9 Citric Acid 0.5 Total weight 263.7

The foamed egg white matrix comprising a functional emulsion was madeaccording to the following process;

-   -   (a) Ingredients 1 (purified water) and 2 (gelatin) from Table 4        were combined in the amounts delineated in a suitable vessel at        room temperature and allowed about 30 minutes to fully hydrate        the gelatin,    -   (b) ingredient 3 was made by adding 40 grams of egg white to 60        grams of water, wetting the powder carefully using a spatula and        allowing it to fully hydrate over a period of about 30 mins,    -   (c) the mixtures from steps (a) and (b) were combined and        blended together,    -   (d) ingredient 4 (hydrogen peroxide) from Table 4 was added in        the amounts delineated to the mixture from step (c) and blended        in well, then ingredient 5 (catalase) from Table 4 was added in        the amounts delineated and blended in well, the foaming reaction        started within one minute and continued for a period of        approximately 5 minutes during which time the foam mass was        stirred continuously,    -   (e) heating was commenced while stirring constantly and using a        suitable thermocouple in the foam and the foam mass was brought        up to 80° C.,    -   (f) ingredient 6 (Sorbitol) from Table 4 was added in the        amounts delineated and stirred in. The foam mass will cool to        approximately 60° C. due to the endothermic solubilization of        the sorbitol,    -   (g) the temperature of the foam mass was held/maintained (using        a water bath or Bain Marie) at 60° C., and    -   (h) ingredient 7 (flavoring) from Table 4 was added in the        amounts delineated and stirred in.

The foam mass can be held at 60° C. for periods of up to 5 hours withoutsignificant deterioration. The foam mass can be dispensed to molds andallowed to cool where it forms a firm flexible material.

Assay of Antimicrobial Effect:

The total weight of the formulation in Example 4 was 263.7 gramscontaining 4 grams of functional emulsion (1.5%). The functionalemulsion contained 10% free Caprylic acid in the oil phase and so theconcentration of free Caprylic acid in the formulation of Example 4 was0.15%.

The Antimicrobial assay as described in the Materials and Methodssection above gave the following result:

Inoculum of Staphylococcus aureus 2.66 × 10⁸ Viable cells per ml, Zerotime in the assay 1.95 × 10⁵ Viable cells per ml, One Minute Exposure2.58 × 10³ Viable cells per ml, and One Minute Log Reduction 1.37 Logs.

Example 5 Egg White/Gelatin/Agar/Chelating Agent Formulation Using anOil in Water Emulsion as a Functional Ingredient

Illustrating the incorporation of sodium hexametaphosphate as achelating agent with amplified citric acid to counteract its intrinsicalkalinity.

TABLE 5 Foamed egg white matrix formulation of Example 5. IngredientWeight in grams 1 Purified water 350 2 Citric Acid monohydrate 2.0 3Agar 7.6 4 Gelatin 260 bloom 40 5 Sorbitol 60 6 Egg white at 20% protein45 concentration whisked to firm peaks 7 Flavoring powder 6 8 Sodiumhexametaphosphate 4 9 Functional emulsion based on 10 U.S. patentapplication 15/384,372 Total weight 520.6

The foamed egg white matrix comprising precursors to a functionalemulsion was made according to the following process;

-   -   (a) Ingredients 1 (purified water), 2 (citric acid monohydrate),        and 3 (agar) from Table 5 were combined in the amounts        delineated in a suitable vessel at room temperature and allowed        about 10 minutes to hydrate, and then heated to about 80° C.        using a thermocouple in the mass to confirm temperature.    -   (b) in a separate container—ingredients 4 (gelatin) and 5        (sorbitol) ingredient 3 (sodium Caprylate) from Table 5 were        combined in the amounts delineated and blended as a dry powder,    -   (c) using suitable scale equipment with whisk attachments        carefully ingredient 6 (20% egg white) from Table 5 in the        amounts delineated was whisked/beaten in a suitable vessel for        approximately 5 minutes to achieve a foam mass that holds its        shape when pulled upwards —firm peaks.    -   (d) When the mixture from step (a) reached a temperature about        80° C. and the agar had melted the powder mixture from step (b)        was added while constantly stirring. The temperature will drop        to approximately 60° C. and step (e) was immediately performed,    -   (e) 45 grams of the mixture from step (c) was added to the        mixture from step (d) and stirred vigorously, and continuously        stirred while heating till the temperature exceeds 80° upon        which heating was turned off and/or the mixture was removed from        heating,    -   (f) ingredient 7 (flavoring) from Table 5 was added in the        amounts delineated and stirred in, then ingredient 8 (sodium        hexametaphosphate) from Table 5 was added in the amounts        delineated and stirred in, them ingredient 9 (functional        emulsion) from Table 5 was added in the amounts delineated and        stirred in and the pH was checked to be below 5.0.

The foam mass can be held at 60° C. for periods of up to 5 hours withoutsignificant deterioration. The foam mass can be dispensed to molds andallowed to cool where it forms a firm flexible material.

In other embodiments sorbitol can be replaced with an equal amount ofErythritol, or another polyol, if required.

Assay of Antimicrobial Effect:

The total weight of the formulation in Example 5 was 520.6 gramscontaining 10 grams of functional emulsion (1.9%).

Using Antimicrobial assay as described in the Materials and Methodssection above the antimicrobial effect was determined to be similar toExample 2 with a 1.6 log reduction in viability after 1 minute ofexposure.

TABLE 6 Summary of Antimicrobial Efficacy. Example Example ExampleExample 1 2 3 4 Inoculum of Staph  1.7 × 10⁹ 2.8 × 10⁹ 1.83 × 10⁹ 2.66 ×10⁸ aureus Viable count at zero 1.35 × 10⁶ 1.9 × 10⁶ 2.54 × 10⁶ 1.95 ×10⁵ time in the assay Viable count at one  2.2 × 10⁴ 2.8 × 10⁴ 1.97 ×10³ 2.58 × 10³ Minute Exposure One Minute Log −1.39 Logs −1.33 Logs −2.1Logs −1.3 Logs Reduction Percent Caprylic 0.14 0.17 0.14 0.15 acidPercent of greatest 66 63 100 62 effect

With the exception of Example 3, while allowing for experimental error,the percentage differences in log reduction in Examples 1, 2, and 4 wasmore or less in line with Caprylic acid content. Notably, Example 3 hadthe same concentration of Caprylic acid as Example 1 and exhibited a 34%greater effect. Although not wishing to be bound by the explanation itis suggested that the increased efficacy in Example 3 compared toExample 1 was due to improved release and available dose at theamplified surface of the whisked egg white due to the more finelydispersed nature of the in situ formation of Caprylic acid, compared toits droplet form when incorporated as an emulsion.

Example 6: Synergistic Antimicrobial Effect of Polyphosphate whenCombined with Functional Emulsion of Free Fatty Acid

The inclusion of hexametaphosphate as an example of a polyphosphate usedas a chelating agent is illustrated in Example 5 wherein sodiumhexametaphosphate (“SHMP” or “sodium HMP”) was included. When evaluatingthe antimicrobial effect of the formulation in Example 5 it wasunexpectedly discovered that the combination of the functional freefatty acid emulsion was acting synergistically with thehexametaphosphate.

All mineral chelating agents will affect a growth suppressing effect onbacteria particularly where essential minerals such as calcium,magnesium, zinc and iron are limited in the media, they achieve thiseffect by sequestration of the essential minerals. The growthsuppressing effect of a chelating agent is easily overcome bysupplementing the growth media with the necessary mineral.

A functional emulsion of free fatty acids as disclosed in U.S. patentapplication Ser. No. 15/384,372 will exert a potent antimicrobial effectas illustrated in Examples 1 to 5 of this application. It wasunexpectedly discovered that the functional emulsion amplified thegrowth suppressing effect of the polyphosphate and did so in thepresence of supplemented calcium which would otherwise suppress theeffect of the hexametaphosphate.

In order to illustrate the synergistic effect of the combination offunctional emulsion and hexametaphosphate it is necessary to constructformulations of both which are at and below their Minimum InhibitoryConcentration and then combine these to measure amplification of effectwhich is greater than what might be expected as additive.

In the present Example a formulation of functional emulsion wasconstructed using 8% W/W Caprylic acid and 2% W/W mixed capric/Caprylictri-glyceride (Miglyol 812N from IOI Oleo, Germany) in the oil phaseusing 1.56% W/W lecithin and 0.15% Tween 80 as co-surfactant usingprocedures disclosed in U.S. patent application Ser. No. 15/384,372.

The test organism is Staphylococcus aureus NCTC 8325-4 as described inthe methods and flask cultures are 100 ml of Luria-Bertani broth (LBbroth) in 250 ml Erlenmeyer flasks. LB broth is used instead of brainHeart Infusion because it is more defined and limited in mineralcontent.

A standard inoculum is grown as described in the methods section andadjusted to a viable cell count of 1×10⁶ by dilution in sterile saline,0.1 ml of this is used to aseptically inoculate 100 ml volumes of testmedia—the inoculum being 1×10³ in the test flask.

The Minimum Inhibitory Concentration of the functional emulsion wasdetermined using the agar dilution method wherein LB agar plates ofprogressively increasing dilution of functional emulsion are streakedwith test organism and evaluated visually for growth after 24 hoursincubation at 37° C.

Results are presented in Table 7

Some growth is still detected at 0.5% and so the Minimum InhibitoryConcentration may be considered to be greater than 0.5%: a concentrationof 0.375 has no inhibitory effect on growth and this concentration wasselected for further study in combination with hexametaphosphate.

TABLE 7 Functional Emulsion Minimum Inhibition ConcentrationStaphylococcus aureus NCTC 8325-4. Emulsion weight % Growth observation1.56 No growth 1.25 No growth 0.937 No growth 0.625 No growth 0.563 Nogrowth 0.5 Trace pin-head colonies 0.438 Few pin-head colonies 0.375Florid growth 0.313 Florid growth 0 Florid growth

To determine the Minimum Inhibitory effect of hexametaphosphate a seriesof flasks with 100 ml of LB broth were each supplemented withconcentrations of hexametaphosphate ranging from 0% W/W to 1% W/W, eachflask was inoculated with a standard inoculum to achieve 1×10³ viablebacterial in the flask, the flasks were incubated at 37° C. in a rotaryincubator for 24 hours and evaluated visually for growth at that time.As illustrated in the first row of Table 8 below growth was evident onlyin the flask with zero concentration of hexametaphosphate, at allconcentrations from 0.05% and above growth was apparently inhibited.

Following visual evaluation, 0.5% sterile calcium chloride was added toall flasks and incubation continued for a further 24 hours. Visualexamination at 24 hours post addition of calcium chloride revealed fullgrowth in all flasks up to and including 0.27% hexametaphosphate.Hexametaphosphate concentrations including and above 0.5%hexametaphosphate remained inhibited. From this data it can be seen that0.27% hexametaphosphate will inhibit growth unless there is mediasupplementation of at least 0.5% calcium chloride in which case there isno inhibitory effect on growth.

TABLE 8 Sodium hexametaphosphate effect on the growth of S. aureus inliquid culture. SHMP % 1.0 0.75 0.5 0.27 0.26 0.25 0.24 0.23 0.22 0.20.15 0.1 0.05 0 Growth at 24 hr − − − − − − − − − − − − − + Add CalciumAll Flasks supplemented with sterile 0.5% Calcium chloride Growth at 24− − − + + + + + + + + + + + hr post CaCl₂ supplementation

Full growth inhibition will be re-instated however if the media isfurther supplemented with 0.375% functional emulsion, a concentrationwhich on its own has no growth inhibitory effect.

The synergistic effect is exemplified in the data presented in Table 9below when supplementing with a functional emulsion which is an oil inwater emulsion consisting of finely dispersed droplets of Caprylic acidoil in water which is stabilized with a purified amphipathic lecithinand optionally other ingredients such as co-surfactants (hereinafterreferred to as “ML:8”).

TABLE 9 ML:8 Hexametaphosphate Synergy. Flask 12 h 24 h 24 H CFU/ml 1Control + + >10⁹ 2 0.375% functional emulsion + + >10⁸ 3 0.5%Calcium + + >10⁹ 4 0.27% SHMP − − ~2 × 10² 5 0.27% SHMP + 0.5% + + >10⁹Calcium 6 0.375% functional emulsion + + >10⁸ plus 0.5% calcium chloride7 0.27% SHMP + 0.5% − − ~2 × 10² Calcium + 0.375% functional emulsion

Row 1 in Table 9 is the control flask containing Luria-Bertani (LB)broth and standard inoculum, the terminal plate count shows full growthwith greater than 10⁹ viable organisms per ml.

Row 2 in Table 9 is the same LB broth with the same inoculumsupplemented with 0.375% functional emulsion, there is florid growth at12 and 24 hours and a terminal plate count showing just one log lessviable cells compared to the control: one log reduction.

Row 3 in Table 9 is a control for calcium chloride showing that thissupplement has no effect on growth with the terminal count the same asthe control.

Row 4 in Table 9 illustrates that 0.27% hexametaphosphate suppressesgrowth entirely with a residual viability of 2×10² which is essentiallythe inoculum that remains viable although suppressed (microbistaticeffect not microbicidal).

Row 5 in Table 9 is the same as Row 4 supplemented with 0.5% calciumchloride showing complete reversal of the inhibitory effect ofhexametaphosphate with terminal viability the same as the control inRow.

Row 6 in Table 9 is the same as Row 2 supplemented with 0.5% calciumchloride, terminal viability is the same as Row 2 with one logreduction—the addition of calcium chloride does amplify or inhibit theeffect of the functional emulsion at the concentrations used in thisExample.

Row 7 in Table 9 is the combination of Rows 2 (no inhibition) and 5 (noinhibition) showing full inhibition with residual viability of just2×10² which is the inhibited inoculum that remains viable throughout thetest procedure.

It can be concluded that under conditions of excess calcium, sodiumhexametaphosphate has no inhibitory effect on microbial growth. Equallyunder similar conditions, 0.375% functional emulsion has no inhibitoryeffect on microbial growth. In combination, in conditions of excesscalcium the two components (functional emulsion and hexametaphosphate)act synergistically to completely inhibit microbial growth.

Similar synergistic effects may be demonstrated using otherpolyphosphates including but not limited to di-sodium orthophosphate,uridine monophosphate and sodium phytate. Using procedures described togenerate the data in Table 8 the critical limits of 0.75% di-sodiumorthophosphate and 0.2% sodium phytate in combination with 0.5% calciumchloride were identified. Uridine monophosphate was inconclusive at 1%,suggesting its chelating properties are less tenacious.

Using similar procedures, the critical limits for non-phosphatechelators including tri-sodium citrate and Ethylenediaminetetraaceticacid in combination with 0.5% calcium chloride are 0.5% and 0.25%respectively.

It should be noted that the ion binding properties of many chelators ispH dependent, particularly in the case of tri-sodium citrate which is anorganic acid salt which dissociates at pH below 6.0.

It will be appreciated by those skilled in the art that the ratios ofchelator/calcium chloride/functional emulsion provided in this examplehave been selected as most appropriate to illustrate the synergisticeffect by comparative measurement. That same synergy is manifest in allcombinations of the same ingredients even though its measurement mightbe occluded by excess of any one ingredient.

Example 7 Alternative Formulations with Functional Emulsion Precursorsin the Egg White Foam

Exemplary formulations of the Foamed egg white matrix have thecompositions disclosed in Table 10.

TABLE 10 Exemplary foamed egg white matrix formulation of Example 7.Ingredient Weight in % 1 Water 55-65 2 Glycerine 13-16 3 Lecithin S75from Lipoid AG 0.05-0.50 4 Sodium Caprylate 0.2-1.0 5 Agar 1.0-2.5 6Sorbitol  5-15 7 Gelatin 2-7 8 Whisked Egg White  2-20 9 Functionalingredient 0-1 10 Sugar 0.1-1.0 11 Flavoring 0.3-2.0 12 Citric acid0.1-1.5

In embodiments the sugar is saccharin.

In embodiments the flavoring is vanillin.

In embodiments the functional ingredient is sodium HMP.

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 11.

TABLE 11 Foamed egg white matrix formulation. Ingredient Weight in grams1 Water 210 2 Glycerine 50 3 Lecithin S75 from Lipoid AG 1 4 SodiumCaprylate 2.32 5 Agar 5 6 Sorbitol 40 7 Gelatin 10 8 Whisked Egg White(20% W/W protein) 10 9 Sodium HMP 1 10 Saccharin 1 11 Vanillin 2 12Citric acid 2 Total weight 334.32

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 12.

TABLE 12 Foamed egg white matrix formulation. Ingredient Weight in grams1 Water 215 2 Glycerine 50 3 Lecithin S75 from Lipoid AG 0.5 4 SodiumCaprylate 1.16 5 Agar 6.5 6 Sorbitol 30 7 Gelatin 20 8 Whisked Egg White(10% W/W protein) 30 9 Sodium HMP 2 10 Saccharin 2 11 Vanillin 4 12Citric acid 3 Total weight 364.16

Example 8 Alternative Formulations with Functional Emulsion Precursorsin the Egg White Foam

Exemplary formulations of the Foamed egg white matrix have thecompositions disclosed in Table 13.

TABLE 13 Exemplary foamed egg white matrix formulation of Example 8Weight in Ingredient grams % 1 Water 40-60 2 Glycerine 13-15 3 Agar1.0-2.5 4 Sorbitol  5-15 5 Gelatine  1-10 6 Whisked Egg White  5-25 7Functional ingredient 0-1 8 Sugar 0.1-1   9 Flavoring 0.3-2.0 10 Citricacid 0.5-7.0 11 Functional emulsion  3-10

In embodiments the sugar is saccharin.

In embodiments the flavoring is vanillin.

In embodiments functional ingredient is sodium HMP. In other embodimentsthere is no addition of a function ingredient other than the functionalemulsion.

In embodiments the functional emulsion is an oil in water emulsionconsisting of finely dispersed droplets of Caprylic acid oil in waterwhich is stabilized with a purified amphipathic lecithin and optionallyother ingredients such as co-surfactants (hereinafter referred to as“ML:8”). In other embodiments the functional emulsion is added asprecursor mixture comprising sodium Caprylate, lecithin, Lipoid S75, andwater (hereinafter referred to as “ML:8 precursor”), which is convertedto the active form of the emulsion after cooking stage using an organicacid to convert the inactive water-soluble sodium Caprylate to theactive oil soluble Caprylic acid.

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 14.

TABLE 14 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 20 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 362.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 15.

TABLE 15 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 20 5 Gelatine 20 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 352.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 16.

TABLE 16 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 10 5 Gelatine 20 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 342.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 17.

TABLE 17 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 15 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 357.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 18.

TABLE 18 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 5 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 347.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 19.

TABLE 19 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 0 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 30 11 ML:8 precursor 18 Total weight 342.5

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 20.

TABLE 20 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 170 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 20 6 Whisked Egg White (10% W/W protein) 30 8Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/W solution) 15 11 ML:8precursor 2.25 Total weight 329.75

In one embodiment the delivery system comprises an egg white matrixformulation as disclosed in Table 21.

TABLE 21 Exemplary foamed egg white matrix formulation of Example 8Weight Ingredient in grams 1 Water 200 2 Glycerine 50 3 Agar 6.5 4Sorbitol 30 5 Gelatine 20 6 Whisked Egg White (10% W/W protein) 30 7Sodium HMP 2 8 Saccharin 2 9 Vanillin 4 10 Citric acid (20% W/Wsolution) 15 11 ML:8 precursor 2.25 Total weight 361.75

1. A delivery system for functional ingredients comprising one or morefunctional ingredient(s) substantially uniformly dispersed in a matrix,wherein said matrix comprises: i) an egg white component comprisingbetween 1-50% protein; ii) one or more heat resistant and/or heatsensitive gelling agents; iii) a pH regulator; iv) one or moreplasticizers and/or humectants; and v) one or more source of water. 2.The delivery system according to claim 1, wherein said one or moregelling agents comprises agar.
 3. The delivery system according to claim1, wherein said one or more gelling agents comprises gelatin.
 4. Thedelivery system according to claim 1, wherein said one or more gellingagents comprises pectin.
 5. The delivery system according to claim 1,wherein said one or more gelling agents comprises at least two gellingagents selected from agar, gelatin, and pectin.
 6. The delivery systemaccording to claim 1, wherein said pH regulator is citric acid.
 7. Thedelivery system according to claim 1, wherein said one or morefunctional ingredient(s) comprises a functional emulsion.
 8. Thedelivery system according to claim 1, comprising two or more functionalingredient(s).
 9. The delivery system according to claim 7, furthercomprising a chelating agent.
 10. The delivery system according to claim9, wherein the chelating agent is a polyphosphate.
 11. The deliverysystem according to claim 10, wherein the polyphosphate is sodiumhexametaphosphate.
 12. The delivery system according to claim 1, whereinsaid one or more functional ingredients are selected from the group ofdrugs, botanicals, nutritional supplements, vitamins, minerals, enzymes,hormones, proteins, polypeptides and antigens.
 13. The delivery systemaccording to claim 1, further comprising a sweetener, a buffer, anatural or artificial flavoring, a coloring agent, or a combinationthereof.
 14. A dual action dental chew for dogs, wherein the dental chewcomprises two components; a flexible base and a filling, and wherein thefilling comprises the delivery system according to claim
 1. 15. The dualaction dental chew for dogs according to claim 14, wherein the flexiblebase is shaped and designed to reduce plaque and calculus throughmechanical action (scrubbing, abrasion) during the chewing process, andwherein the chew base features a ‘reservoir’/cavity for the filling. 16.The dual action dental chew for dogs according to claim 14, wherein thedelivery system comprises a functional ingredient comprising an emulsioncomprising (a) one or more saturated or unsaturated free fatty acidshaving from 4 to 22 carbon atoms or a pharmaceutically acceptable saltthereof; and (b) one or more delipidised membrane lipids, as emulsifyingagent for the free fatty acid(s) or the salt thereof.
 17. A method ofusing the delivery system according to claim 1 for oral administrationof one or more functional ingredient to an animal in need thereofcomprising administering the delivery system to the animal in need.